DNA Barcoding of Black Flies (Diptera: Simuliidae) in Slovakia and Its Utility for Species Identification

DNA barcoding based on the cytochrome oxidase I gene is increasingly used in black flies (Diptera: Simuliidae), but extensive data for larger areas are still rare. Slovakia, with well-explored black fly fauna, was chosen to verify the reliability of DNA barcoding for species identification. The DNA barcoding region of the COI gene of 235 individuals of 25 black fly species from Slovakia was sequenced. Among them, 30 sequence clusters with assigned Barcode Index Numbers (BINs) were identified, and 5 of them were recorded for the first time. The average intraspecific genetic divergence was 0-3.24%, whereas the average interspecific divergence was 12.3-17.8%. Based on the barcode sequence, 14 species could be identified unambiguously, and 3 of them (Prosimulium latimucro, Simulium costatum, S. degrangei) are split into two or more barcode clusters. In eleven species, some degree of barcode sharing occurred, often combined with barcode splitting. The results showed hidden diversity as well as cases of shared barcode sequences among the studied species. Further investigation using other molecular markers is necessary due to the overlap of intraspecific and interspecific variability.